The complexity of eukaryotic proteomes—that is, the total number of distinct protein species present concurrently in a eukaryotic cell, including alternatively spliced isoforms and variants differing in post-translational modification—typically exceeds the resolving capacity of current analytical techniques.
For example, the number of distinct protein species in eukaryotic cells typically far exceeds the spatial resolution of two-dimensional polyacrylamide gel electrophoresis (2D PAGE) gels, with large numbers of distinct protein species appearing to comigrate. The limited spatial resolution in turn constrains the dynamic detection range of the technique: efforts to observe low abundance species by increasing the initial protein load lead to increased obscuration by high abundance species.
Recently, efforts have been made to increase resolution of such protein analytical techniques by prefractionating the protein mixture prior to analysis. In one approach, complex mixtures are prefractionated using solution phase isoelectric focusing, yielding fractions having distinct pI ranges that can thereafter be separately analyzed by 2D PAGE with increased resolution. See Zuo and Speicher, Anal. Biochem. 284:266-278 (2000); Zuo et al., Electrophoresis 22:1603-1615 (2001); Zuo and Speicher, Proteomics 2:58-68 (2002); Ali-Khan et al., Current Protocols in Protein Science 22.1:1-19 (2002); Zuo et al., J. Chromatography B 782:253-265 (2002); and Wistar Institute, WO 01/75432, the disclosures of which are incorporated herein by reference in their entireties. See also Tan et al., Electrophoresis 23:3599-3607 (2002); WO 01/36449; WO 00/17631; and Righetti et al., J. Chromatography 475:293-309 (1989).
A number of devices capable of solution phase isoelectric focusing are available commercially. None of the devices, however, provides particularly convenient solution phase IEF prefractionation of small volume protein samples with a simple device in a format that readily interfaces with subsequent analytical techniques such as 2D PAGE.
There thus exists a continuing need in the art for devices, components, and methods for solution phase electrophoresis, particularly solution phase isoelectric focusing.